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Highly multiplexed and quantitative cell-surface protein profiling using genetically barcoded antibodies.

  • Author(s): Pollock, Samuel B
  • Hu, Amy
  • Mou, Yun
  • Martinko, Alexander J
  • Julien, Olivier
  • Hornsby, Michael
  • Ploder, Lynda
  • Adams, Jarrett J
  • Geng, Huimin
  • Müschen, Markus
  • Sidhu, Sachdev S
  • Moffat, Jason
  • Wells, James A
  • et al.
Abstract

Human cells express thousands of different surface proteins that can be used for cell classification, or to distinguish healthy and disease conditions. A method capable of profiling a substantial fraction of the surface proteome simultaneously and inexpensively would enable more accurate and complete classification of cell states. We present a highly multiplexed and quantitative surface proteomic method using genetically barcoded antibodies called phage-antibody next-generation sequencing (PhaNGS). Using 144 preselected antibodies displayed on filamentous phage (Fab-phage) against 44 receptor targets, we assess changes in B cell surface proteins after the development of drug resistance in a patient with acute lymphoblastic leukemia (ALL) and in adaptation to oncogene expression in a Myc-inducible Burkitt lymphoma model. We further show PhaNGS can be applied at the single-cell level. Our results reveal that a common set of proteins including FLT3, NCR3LG1, and ROR1 dominate the response to similar oncogenic perturbations in B cells. Linking high-affinity, selective, genetically encoded binders to NGS enables direct and highly multiplexed protein detection, comparable to RNA-sequencing for mRNA. PhaNGS has the potential to profile a substantial fraction of the surface proteome simultaneously and inexpensively to enable more accurate and complete classification of cell states.

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